In normal heart development a primitive heart tube forms along the midline. This relatively straight tube bends and rotates towards the right, initiating the morphogenesis of the organ. Reverse bending (dextro-cardia) occurs in only 0.011% of the adult human population, associated with a very high frequency of congenital heart disease. The asymmetry of the heart is the first morphological manifestation of the left-right (LR) asymmetry during embryogenesis. The investigators have recently identified in chick the first three examples of genes expressed asymmetrically on the left and right sides prior to morphological asymmetry. They each encode secreted proteins of the activin, hedgehog and nodal gene families. These genes, expressed during and after gastrulation, regulate each other's expression in a sequential pathway. Moreover, manipulation of the sidedness of their expression alters heart situs. This discovery puts the investigators in a unique position to dissect the molecular and cellular regulation of LR heart asymmetry. They have already demonstrated that activin and sonic hedgehog regulate heart situs. The third member of the LR signalling cascade, cNR-1 (related to the mouse gene nodal) will be expressed in cells in culture, and implanted into embryos to assess its effect on heart laterality. The involvement of this pathway in establishing asymmetry of other organs will also be assessed. Twin chick embryos will be constructed to determine whether cross-embryonic signaling is responsible for the clinical observation of situs inversus in conjoined twins. Manipulation of these signals will allow the current models of heart tube bending to be tested, including related contributions to the caudal and rostral heart from the left and right cardiac primordia and organization of actin filaments. The murine homologue of cNR-1 will be cloned and its expression will be examined in mutants which exhibit reversed heart situs. In collaboration with Andy McMahon's laboratory, mNR-1 will be genetically mapped and the mNR-1 gene will be knocked out.